scholarly journals A Replicase of Potato virus X Acts as the Resistance-Breaking Determinant for JAX1-Mediated Resistance

2013 ◽  
Vol 26 (9) ◽  
pp. 1106-1112 ◽  
Author(s):  
Kyoko Sugawara ◽  
Takuya Shiraishi ◽  
Tetsuya Yoshida ◽  
Naoko Fujita ◽  
Osamu Netsu ◽  
...  
Keyword(s):  
Amino Acid ◽  
Potato Virus ◽  
Potato Virus X ◽  
Single Amino Acid ◽  
Wild Type ◽  
Methyl Transferase ◽  
General Role ◽  
Resistance Breaking ◽  
Viral Replicase

Lectin-mediated resistance (LMR) has been suggested to comprise an uncharacterized branch of antiviral plant innate immunity. To unveil the feature of resistance conferred by jacalin-type lectin required for potexvirus resistance 1 (JAX1), a recently isolated LMR gene against potexviruses, we analyzed the resistance-breaking variants to find the viral component involved in resistance. We employed grafting-mediated inoculation, a high-pressure virus inoculation method, to obtain Potato virus X (PVX) variants that can overcome JAX1-mediated resistance. Whole-genome sequencing of the variants suggested that a single amino acid in the methyl transferase domain of the replicase encoded by PVX is responsible for this resistance-breaking property. Reintroduction of the amino-acid substitution to avirulent wild-type PVX was sufficient to overcome the JAX1-mediated resistance. These results suggest that viral replicase is involved in JAX1-mediated resistance. The residue that determines the resistance-breaking properties was highly conserved among potexviruses, suggesting a general role of the residue in potexvirus–JAX1 interactions.

A single amino acid residue of RNA-dependent RNA polymerase in the Potato virus X genome determines the symptoms in Nicotiana plants

2005 ◽  
Vol 110 (1-2) ◽  
pp. 177-182 ◽  
Author(s):  
Satoshi Kagiwada ◽  
Yasuyuki Yamaji ◽  
Ken Komatsu ◽  
Shuichiro Takahashi ◽  
Takuma Mori ◽  
...  
Keyword(s):  
Amino Acid ◽  
Rna Polymerase ◽  
Amino Acid Residue ◽  
Potato Virus ◽  
Potato Virus X ◽  
Single Amino Acid ◽  
Rna Dependent Rna Polymerase ◽  
Single Amino Acid Residue

scholarly journals A Single Amino Acid Change in Viral Genome-Associated Protein of Potato Virus Y Correlates with Resistance Breaking in ‘Virgin A Mutant’ Tobacco

1999 ◽  
Vol 89 (2) ◽  
pp. 118-123 ◽  
Author(s):  
Chikara Masuta ◽  
Mitsuyo Nishimura ◽  
Hiroshi Morishita ◽  
Tatsuji Hataya
Keyword(s):  
Amino Acid ◽  
Potato Virus ◽  
Viral Genome ◽  
Potato Virus Y ◽  
Cell Movement ◽  
Amino Acid Sequences ◽  
Single Amino Acid ◽  
Viral Gene ◽  
Resistance Breaking ◽  
Single Amino Acid Change

Tobacco cultivar Virgin A Mutant (VAM) is reported to have the recessive potyvirus resistance gene va. Varied levels of resistance were observed in VAM plants inoculated with Japanese potato virus Y (PVY) isolates. VAM was highly resistant to most of the PVY isolates tested and tolerant to three necrotic strain isolates of PVY-T. Based on data obtained from tissue printing and press blotting, the resistance appeared to be mainly at the level of cell-to-cell movement. PVY replicated in VAM proto-plasts, but the replication was 30% lower than in susceptible tobacco, suggesting that impairment of replication also contributes to resistance. To identify the viral gene product or products involved in VAM resistance, we isolated spontaneous resistance-breaking mutants by passing vein-banding (O strain) isolates several times through VAM plants. By comparing the amino acid sequences of the mutants with their original isolates, we identified a single amino acid substitution in the viral genome-associated protein (VPg) domain that is correlated with VAM resistance breaking. Together, these results suggest that, in addition to its role in replication, VPg plays an important role in the cell-to-cell movement of PVY.

scholarly journals Sensitivity of human immunodeficiency virus to bicyclam derivatives is influenced by the three-dimensional structure of gp120.

1997 ◽  
Vol 41 (12) ◽  
pp. 2616-2620 ◽  
Author(s):  
K De Vreese ◽  
I Van Nerum ◽  
K Vermeire ◽  
J Anné ◽  
E De Clercq
Keyword(s):  
Human Immunodeficiency Virus ◽  
Amino Acid ◽  
Single Amino Acid ◽  
Resistance Pattern ◽  
V3 Loop ◽  
Resistant Virus ◽  
Wild Type ◽  
Immunodeficiency Virus ◽  
Resistant Strains ◽  
Anti Hiv

The bicyclams are a new class of anti-human immunodeficiency virus (anti-HIV) compounds targeted at viral entry. From marker rescue experiments, it appears that the envelope gp120 glycoprotein plays an important role in the anti-HIV activity of the bicyclams. Bicyclam-resistant strains contain a number of amino acid changes scattered over the V2 to V5 region of gp120. Experiments aimed at estimating the relative importance of particular amino acid changes with regard to the overall resistance pattern are described. The sequences of some partially bicyclam-resistant virus strains, obtained during the resistance development process, were analyzed, and the corresponding 50% effective concentrations were determined. Selected mutations observed in bicyclam-resistant strains were introduced in the wild-type background by site-directed mutagenesis. In addition, some amino acids were back-mutated to their wild-type counterparts in an otherwise JM3100-resistant strain. The sensitivities of these mutant viruses to bicyclams were determined. Construction of chimeric viruses, carrying the V3 loop of JM3100-resistant virus in a wild-type HIV type 1 HXB2 background, enabled us to investigate the importance of the mutations in the V3 loop of JM3100-resistant virus. From the results described in the report, it can be concluded that single amino acid substitutions do not influence the observed resistance to JM3100. Also, the mutations in the V3 loop are not sufficient to engender even a partially resistant phenotype. We postulate that the overall conformation of gp120 determines the degree of sensitivity or resistance of HIV strains to bicyclams.

scholarly journals The Role of Arg13 in Protein Phosphatase M tPphA from Thermosynechococcus elongatus

2012 ◽  
Vol 2012 ◽  
pp. 1-7 ◽  
Author(s):  
Jiyong Su ◽  
Karl Forchhammer
Keyword(s):  
Amino Acid ◽  
Protein Phosphatase ◽  
Arginine Residue ◽  
Side Chain ◽  
Wild Type ◽  
Catalytic Center ◽  
Nitrophenyl Phosphate ◽  
Reduced Activity ◽  
Amino Acid Variants

A highly conserved arginine residue is close to the catalytic center of PPM/PP2C-type protein phosphatases. Different crystal structures of PPM/PP2C homologues revealed that the guanidinium side chain of this arginine residue can adopt variable conformations and may bind ligands, suggesting an important role of this residue during catalysis. In this paper, we randomly mutated Arginine 13 of tPphA, a PPM/PP2C-type phosphatase from Thermosynechococcus elongatus, and obtained 18 different amino acid variants. The generated variants were tested towards p-nitrophenyl phosphate and various phosphopeptides. Towards p-nitrophenyl phosphate as substrate, twelve variants showed 3–7 times higher Km values than wild-type tPphA and four variants (R13D, R13F, R13L, and R13W) completely lost activity. Strikingly, these variants were still able to dephosphorylate phosphopeptides, although with strongly reduced activity. The specific inability of some Arg-13 variants to hydrolyze p-nitrophenyl phosphate highlights the importance of additional substrate interactions apart from the substrate phosphate for catalysis. The properties of the R13 variants indicate that this residue assists in substrate binding.

Biochemical and genetic characterization of a yeast TFIID mutant that alters transcription in vivo and DNA binding in vitro

1992 ◽  
Vol 12 (5) ◽  
pp. 2372-2382
Author(s):  
K M Arndt ◽  
S L Ricupero ◽  
D M Eisenmann ◽  
F Winston
Keyword(s):  
Amino Acid ◽  
Dna Binding ◽  
Direct Repeat ◽  
Single Amino Acid ◽  
Wild Type ◽  
Dna Binding Specificity ◽  
Selection For

A mutation in the gene that encodes Saccharomyces cerevisiae TFIID (SPT15), which was isolated in a selection for mutations that alter transcription in vivo, changes a single amino acid in a highly conserved region of the second direct repeat in TFIID. Among eight independent spt15 mutations, seven cause this same amino acid change, Leu-205 to Phe. The mutant TFIID protein (L205F) binds with greater affinity than that of wild-type TFIID to at least two nonconsensus TATA sites in vitro, showing that the mutant protein has altered DNA binding specificity. Site-directed mutations that change Leu-205 to five different amino acids cause five different phenotypes, demonstrating the importance of this amino acid in vivo. Virtually identical phenotypes were observed when the same amino acid changes were made at the analogous position, Leu-114, in the first repeat of TFIID. Analysis of these mutations and additional mutations in the most conserved regions of the repeats, in conjunction with our DNA binding results, suggests that these regions of the repeats play equivalent roles in TFIID function, possibly in TATA box recognition.

Single amino acid substitution of rat MRP2 results in acquired transport activity for taurocholate

2001 ◽  
Vol 281 (4) ◽  
pp. G1034-G1043 ◽  
Author(s):  
Kousei Ito ◽  
Hiroshi Suzuki ◽  
Yuichi Sugiyama
Keyword(s):  
Amino Acids ◽  
Multidrug Resistance ◽  
Amino Acid ◽  
Membrane Vesicles ◽  
Single Amino Acid ◽  
Sf9 Cells ◽  
Transport Activity ◽  
Wild Type ◽  
Cationic Amino Acids ◽  
The Difference

Multidrug resistance-associated protein 3 (MRP3), unlike other MRPs, transports taurocholate (TC). The difference in TC transport activity between rat MRP2 and MRP3 was studied, focusing on the cationic amino acids in the transmembrane domains. For analysis, transport into membrane vesicles from Sf9 cells expressing wild-type and mutated MRP2 was examined. Substitution of Arg at position 586 with Leu and Ile and substitution of Arg at position 1096 with Lys, Leu, and Met resulted in the acquisition of TC transport activity, while retaining transport activity for glutathione and glucuronide conjugates. Substitution of Leu at position 1084 of rat MRP3 (which corresponds to Arg-1096 in rat MRP2) with Lys, but not with Val or Met, resulted in the loss of transport activity for TC and glucuronide conjugates. These results suggest that the presence of the cationic charge at Arg-586 and Arg-1096 in rat MRP2 prevents the transport of TC, whereas the presence of neutral amino acids at the corresponding position of rat MRP3 is required for the transport of substrates.

scholarly journals Mutations in Potato virus Y Genome-Linked Protein Determine Virulence Toward Recessive Resistances in Capsicum annuum and Lycopersicon hirsutum

2004 ◽  
Vol 17 (3) ◽  
pp. 322-329 ◽  
Author(s):  
Benoît Moury ◽  
Caroline Morel ◽  
Elisabeth Johansen ◽  
Laurent Guilbaud ◽  
Sylvie Souche ◽  
...  
Keyword(s):  
Amino Acid ◽  
Capsicum Annuum ◽  
Potato Virus ◽  
Potato Virus Y ◽  
Cdna Clones ◽  
Lycopersicon Hirsutum ◽  
Potato Virus A ◽  
Single Nucleotide Change ◽  
Protein Encoding

The recessive resistance genes pot-1 and pvr2 in Lycopersicon hirsutum and Capsicum annuum, respectively, control Potato virus Y (PVY) accumulation in the inoculated leaves. Infectious cDNA molecules from two PVY isolates differing in their virulence toward these resistances were obtained using two different strategies. Chimeras constructed with these cDNA clones showed that a single nucleotide change corresponding to an amino acid substitution (Arg119His) in the central part of the viral protein genome-linked (VPg) was involved in virulence toward the pot-1 resistance. On the other hand, 15 nucleotide changes corresponding to five putative amino acid differences in the same region of the VPg affected virulence toward the pvr21 and pvr22 resistances. Substitution models identified six and five codons within the central and C terminal parts of the VPg for PVY and for the related potyvirus Potato virus A, respectively, which undergo positive selection. This suggests that the role of the VPg-encoding region is determined by the protein and not by the viral RNA apart from its protein-encoding capacity.

scholarly journals A Single Amino Acid Substitution in the West Nile Virus Nonstructural Protein NS2A Disables Its Ability To Inhibit Alpha/Beta Interferon Induction and Attenuates Virus Virulence in Mice

2006 ◽  
Vol 80 (5) ◽  
pp. 2396-2404 ◽  
Author(s):  
Wen Jun Liu ◽  
Xiang Ju Wang ◽  
David C. Clark ◽  
Mario Lobigs ◽  
Roy A. Hall ◽  
...  
Keyword(s):  
West Nile Virus ◽  
Amino Acid ◽  
Amino Acid Substitution ◽  
Nonstructural Protein ◽  
Mutant Virus ◽  
Single Amino Acid Substitution ◽  
Single Amino Acid ◽  
Wild Type ◽  
Alpha Beta ◽  
A30p Mutation

ABSTRACT Alpha/beta interferons (IFN-α/β) are key mediators of the innate immune response against viral infection. The ability of viruses to circumvent IFN-α/β responses plays a crucial role in determining the outcome of infection. In a previous study using subgenomic replicons of the Kunjin subtype of West Nile virus (WNVKUN), we demonstrated that the nonstructural protein NS2A is a major inhibitor of IFN-β promoter-driven transcription and that a single amino acid substitution in NS2A (Ala30 to Pro [A30P]) dramatically reduced its inhibitory effect (W. J. Liu, H. B. Chen, X. J. Wang, H. Huang, and A. A. Khromykh, J. Virol. 78:12225-12235). Here we show that incorporation of the A30P mutation into the WNVKUN genome results in a mutant virus which elicits more rapid induction and higher levels of synthesis of IFN-α/β in infected human A549 cells than that detected following wild-type WNVKUN infection. Consequently, replication of the WNVKUNNS2A/A30P mutant virus in these cells known to be high producers of IFN-α/β was abortive. In contrast, both the mutant and the wild-type WNVKUN produced similar-size plaques and replicated with similar efficiency in BHK cells which are known to be deficient in IFN-α/β production. The mutant virus was highly attenuated in neuroinvasiveness and also attenuated in neurovirulence in 3-week-old mice. Surprisingly, the mutant virus was also partially attenuated in IFN-α/βγ receptor knockout mice, suggesting that the A30P mutation may also play a role in more efficient activation of other antiviral pathways in addition to the IFN response. Immunization of wild-type mice with the mutant virus resulted in induction of an antibody response of similar magnitude to that observed in mice immunized with wild-type WNVKUN and gave complete protection against challenge with a lethal dose of the highly virulent New York 99 strain of WNV. The results confirm and extend our previous original findings on the role of the flavivirus NS2A protein in inhibition of a host antiviral response and demonstrate that the targeted disabling of a viral mechanism for evading the IFN response can be applied to the development of live attenuated flavivirus vaccine candidates.

scholarly journals Mechanism of Darunavir (DRV)’s High Genetic Barrier to HIV-1 Resistance: A Key V32I Substitution in Protease Rarely Occurs, but Once It Occurs, It Predisposes HIV-1 To Develop DRV Resistance

mBio ◽  
2018 ◽  
Vol 9 (2) ◽  
Author(s):  
Manabu Aoki ◽  
Debananda Das ◽  
Hironori Hayashi ◽  
Hiromi Aoki-Ogata ◽  
Yuki Takamatsu ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Amino Acid ◽  
Critical Role ◽  
Viral Fitness ◽  
Single Amino Acid ◽  
Amino Acid Substitutions ◽  
Wild Type ◽  
Genetic Barrier ◽  
High Level ◽  
Hiv 1

ABSTRACTDarunavir (DRV) has bimodal activity against HIV-1 protease, enzymatic inhibition and protease dimerization inhibition, and has an extremely high genetic barrier against development of drug resistance. We previously generated a highly DRV-resistant HIV-1 variant (HIVDRVRP51). We also reported that four amino acid substitutions (V32I, L33F, I54M, and I84V) identified in the protease of HIVDRVRP51are largely responsible for its high-level resistance to DRV. Here, we attempted to elucidate the role of each of the four amino acid substitutions in the development of DRV resistance. We found that V32I is a key substitution, which rarely occurs, but once it occurs, it predisposes HIV-1 to develop high-level DRV resistance. When two infectious recombinant HIV-1 clones carrying I54M and I84V (rHIVI54Mand rHIVI84V, respectively) were selected in the presence of DRV, V32I emerged, and the virus rapidly developed high-level DRV resistance. rHIVV32Ialso developed high-level DRV resistance. However, wild-type HIVNL4-3(rHIVWT) failed to acquire V32I and did not develop DRV resistance. Compared to rHIVWT, rHIVV32Iwas highly susceptible to DRV and had significantly reduced fitness, explaining why V32I did not emerge upon selection of rHIVWTwith DRV. When the only substitution is at residue 32, structural analysis revealed much stronger van der Waals interactions between DRV and I-32 than between DRV and V-32. These results suggest that V32I is a critical amino acid substitution in multiple pathways toward HIV-1’s DRV resistance development and elucidate, at least in part, a mechanism of DRV’s high genetic barrier to development of drug resistance. The results also show that attention should be paid to the initiation or continuation of DRV-containing regimens in people with HIV-1 containing the V32I substitution.IMPORTANCEDarunavir (DRV) is the only protease inhibitor (PI) recommended as a first-line therapeutic and represents the most widely used PI for treating HIV-1-infected individuals. DRV possesses a high genetic barrier to development of HIV-1’s drug resistance. However, the mechanism(s) of the DRV’s high genetic barrier remains unclear. Here, we show that the preexistence of certain single amino acid substitutions such as V32I, I54M, A71V, and I84V in HIV-1 protease facilitates the development of high-level DRV resistance. Interestingly, allin vitro-selected highly DRV-resistant HIV-1 variants acquired V32I but never emerged in wild-type HIV (HIVWT), and V32I itself rendered HIV-1 more sensitive to DRV and reduced viral fitness compared to HIVWT, strongly suggesting that the emergence of V32I plays a critical role in the development of HIV-1’s resistance to DRV. Our results would be of benefit in the treatment of HIV-1-infected patients receiving DRV-containing regimens.

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